This R21 application proposes a detailed molecular analysis of bacterial isolates recovered as part of the SCOUT Study (Short Course Therapy for Urinary Tract Infections in Children). The parent study, one of the Targeted Clinical Trials to Reduce the Risk of Antimicrobial Resistance, is a multicenter, randomized, double-blind, placebo-controlled non-inferiority clinical trial of children with a confirmed diagnosis of urinary tract infection (UTI). Subjects will be randomized to receive either standard (10-day) or short-course (5-day) therapy with one of three antibiotic regimens: trimethoprim-sulfamethoxazole (TMP-SMX); amoxicillin-clavulanate (AMC); or cefixime (CFX)/cefdinir (CFD). The primary objective of the study is to determine whether short- course therapy is as effective as standard course therapy for treatment of UTI in children; a secondary objective seeks to determine whether the two therapy arms result in similar proportions of children with gastrointestinal colonization by antibiotic-resistant Escherichia coli and/or Klebsiella pneumoniae. Thus, subjects will have stool samples obtained for bacterial culture at enrollment visit (#1; day 4-6) and two follow-up visits, on day 12-14 (#2) and day 24- 26 (#3). In this proposal, we hypothesize that both the antibiotic agent and the length of therapy will affect the patients' indigenous flora and thus affect both the likelihood of recovering E. coli and/or K. pneumoniae from stool and the resistance phenotypes of the recovered isolates. We will utilize PCR- and sequence-based methods to characterize phylogenetic and resistance properties of the recovered isolates. We hypothesize that (1) E. coli and K. pneumoniae recovered from 2 or 3 stool cultures are more likely to be members of disease-associated subgroups within their respective species (E. coli phylogroups B2 and D, K. pneumoniae cluster KpI); (2) treatment-susceptible strains recovered from cultures during treatment (culture #1 for 5-day arm; cultures #1 or #2, 10-day arm) are more likely to be members of disease-associated subgroups; (3A) treatment-resistant strains of either species are more likely than treatment- susceptible strains to be recovered from cultures during treatment; and (3B) treatment- susceptible strains of either species are more likely to be recovered from cultures during treatment in the AMC arm than in the TMP/SMX or CFX/CFD arms. If SCOUT demonstrates that short-course therapy is as safe and effective as standard therapy, and this proposal demonstrates that short-course therapy is less likely to select for intestinal carriage of resistant E. coli or K. pneumoniae, these findings would make a significant impact on prescribing behavior, toward reducing pediatric antibiotic exposure and thus the selection for resistance. PUBLIC HEALTH RELEVANCE: Urinary tract infection (UTI) is one of the most common bacterial infections in pediatric patients, leading to enormous numbers of prescriptions for antibiotic therapy. At the same time, antibiotic resistance in bacteria is on the rise worldwide, primarily due to overuse of antibiotics. This study will determine whether shorter antibiotic treatments for UTI can reduce the selection for carriage of intestinal bacteria resistant to antibiotics.